1. Receptor-mediated endocytosis is an important mechanism for transport of
macromolecules and regulation of cell-surface receptor expression. In rena
l proximal tubules, receptor-mediated endocytosis mediates the reabsorption
of filtered albumin. acidification of the endocytic compartments is essent
ial because it interferes with ligand-receptor dissociation, vesicle traffi
cking, fusion events and coat formation.
2. Here we show that the activity of Na+-H+ exchanger isoform 3 (NHE3) is i
mportant for proper receptor-mediated endocytosis of albumin and endosomal
pH homeostasis in a renal proximal tubular cell line (opossum kidney cells)
which expresses NHE3 only
3. Depending on their inhibitory potency with respect to NHE3 and their lip
ophilicity, the NHE inhibitors EIPA, amiloride and HOE694 differentially re
duced albumin endocytosis. The hydrophilic inhibitor HOE642 had no effect.
4. Inhibition of NHE3 led to an alkalinization of early endosomes and to an
acidification of the cytoplasm, indicating that Na+-H+ exchange contribute
s to the acidification of the early endosomal compartment due to the existe
nce of a sufficient Na+ gradient across the endosomal membrane.
5. Exclusive acidification of the cytoplasm with propionic acid or by remov
al of Na+ induced a significantly smaller reduction in endocytosis than tha
t induced by inhibition of Na+-H+ exchange.
6. Analysis of the inhibitory profiles indicates that in early endosomes an
d endocytic vesicles NHE3 is of major importance, whereas plasma membrane N
HE3 plays a minor role.
7. Thus, NHE3-mediated acidification along the first part of the endocytic
pathway plays an important role in receptor-mediated endocytosis. Furthermo
re, the involvement of NHE3 offers new ways to explain the regulation of re